The state of the art provides methods for producing antibodies in vitro (eg, using phage, ribosome or yeast display) or in vivo (eg, using non-human vertebrates (eg, mice and rats) and cells comprising transgenic immunoglobulin loci). Such in vivo systems (eg, Xenomouse™) have used completely human transgenic heavy chain loci which comprise human variable regions (human VH, D and JH gene segments) upstream of human constant regions (eg, human mu upstream of human gamma constant gene segments). Subsequently, it has been discovered that the use of totally human transgenic loci in such in vivo systems is detrimental and B-cell development is hampered, leading to relatively small B-cell compartments and restricted utility for generating antibodies. Later-generation transgenic animals (eg, the Velocimouse™) have been created which have chimaeric heavy chain loci in which a human variable region is upstream of endogenous (eg, mouse or rat) constant regions (ie, mouse mu constant region upstream of gamma constant region, in germline configuration). This enables the harnessing of endogenous control mechanisms for B-cell and antibody development, and as such the extent of problems of totally human transgenic loci are not seen. Methods of constructing transgenic vertebrates and use of these to generate antibodies and nucleic acids thereof following antigen immunisation are known in the art, eg, see U.S. Pat. No. 7,501,552 (Medarex); U.S. Pat. Nos. 5,939,598 & 6,130,364 (Abgenix); WO02066630, WO2011163311 & WO2011163314 (Regeneron); WO2011004192 & WO2011158009 (Kymab Limited); WO2009076464, WO2009143472, EP1414858, WO2009013620A2, WO2010070263A1 & WO2010109165A2 (Harbour Antibodies); EP1399559 (Crescendo Biologics) and WO2010039900 (Ablexis), the disclosures of which are explicitly incorporated herein including, but not limited to, for the purpose of providing the skilled person with guidance of how to make non-human animals bearing transgenic immunoglobulin loci and to inactivate endogenous loci expression. US2008/0196112A1 (Innate Pharma) discloses transgenic animals comprising a single, predetermined human rearranged VDJ from a lead antibody, together with one or more human constant region genes in a locus. There are no repertoires of unrearranged V, D and J and recombination and junctional diversity to produce a repertoire of VH domains and H chains and antibodies is not addressed.
Whether selected and produced in vitro or in vivo, the art has recognised that the human therapeutic utility of chimaeric antibodies is hampered by their non-human constant domains, despite having human variable domains. This is due to the immunogenicity of the non-human portions once the antibodies have been administered to human patients. The solution in the art to this issue has been to humanise chimaeric antibodies in vitro using protein engineering whereby the non-human constant domains are replaced with corresponding human constant domains. While addressing the immunogenicity issue, however, such engineering in vitro can downgrade the desirable characteristics of the resultant antibody. For example, antigen-binding affinity, antigen specificity, expressibility (eg, in cell lines such as CHO or HEK293 cells), half-life and/or biophysical characteristics (eg, melting temperature, solution state, resistance to aggregation etc) can be downgraded, thereby hampering development of the antibody as drugs for human therapeutic or prophylactic use. It would be desirable to have means for selecting and humanising antibodies that addresses these shortcomings in the art as well as providing repertoires of such humanised antibodies or heavy chains from which drug candidates can be selected.